Panasyuk Ganna, Nemazanyy Ivan, Filonenko Valeriy, Gout Ivan
Institute of Molecular Biology and Genetics, National Academy of Sciences of Ukraine, Kyiv, Ukraine.
Biochem Biophys Res Commun. 2008 May 2;369(2):339-43. doi: 10.1016/j.bbrc.2008.02.016. Epub 2008 Feb 13.
Ribosomal protein S6 kinase (S6K) is involved in the regulation of cell growth and cellular metabolism. The activation of S6K in response to diverse extracellular stimuli is mediated by multiple phosphorylations coordinated by the mTOR and PI3K signaling pathways. We have recently found that both forms of S6K are modified by ubiquitination. Following these findings, we demonstrate here for the first time that S6K1 associates specifically with ubiquitin ligase ROC1 in vitro and in vivo. The interaction was initially identified in the yeast two-hybrid screening and further confirmed by pull-down and co-immunoprecipitation assays. Furthermore, the overexpression of ROC1 leads to an increase in S6K1 ubiquitination. Consistent with this observation, we showed that the steady-state level of S6K1 is regulated by ROC1, since downregulation of ROC1 by specific siRNA promotes stabilization of S6K1 protein. The results suggest the involvement of ROC1 in S6K1 ubiquitination and subsequent proteasomal degradation.
核糖体蛋白S6激酶（S6K）参与细胞生长和细胞代谢的调控。S6K对多种细胞外刺激的激活是由mTOR和PI3K信号通路协调的多重磷酸化介导的。我们最近发现两种形式的S6K都被泛素化修饰。基于这些发现，我们在此首次证明S6K1在体外和体内都与泛素连接酶ROC1特异性结合。这种相互作用最初是在酵母双杂交筛选中鉴定出来的，并通过下拉和免疫共沉淀实验进一步得到证实。此外，ROC1的过表达导致S6K1泛素化增加。与这一观察结果一致，我们表明S6K1的稳态水平受ROC1调控，因为通过特异性小干扰RNA（siRNA）下调ROC1可促进S6K1蛋白的稳定。这些结果表明ROC1参与了S6K1的泛素化及随后的蛋白酶体降解过程。
